Dataset Information

Molecular Disruption of Ion Transport Peptide Receptor Results in Impaired Water Homeostasis and Developmental Defects in Bombyx mori.

ABSTRACT: Insect ion transport peptides (ITPs) are important regulators of many physiological processes and they exert their functions by interacting with their receptors (ITPRs). In the current study, we comprehensively investigated the physiological functions of ITPR in the lepidopteran model insect, the silkworm (Bombyx mori), using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 nuclease (Cas9) genome editing technique. Mutations in silkworm ITPR (BNGR-A2) resulted in a prolongnation of the larval stage by 3.5-day as well as failure in wing expansion of moths. The BNGR-A2 mutation accelerated food transition throughout the digestive tract, which is 1.55-fold that of wild type (WT) insects. Excretion was 1.56-fold of WT insects during the larval stage, resulting in the loss of body water content. Loss of BNGR-A2 function induced significant upregulation of nitric oxide synthase (NOS) enzyme activity and nitric oxide (NO) content, as well as downstream Ca²⁺/NO/cGMP signaling pathways. Key genes in insulin and ecdysone signaling pathways were also affected by BNGR-A2 disruption. Our data show that ITPR plays key roles in regulating insect water homeostasis and development.

SUBMITTER: Sun L

PROVIDER: S-EPMC7251169 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Molecular Disruption of Ion Transport Peptide Receptor Results in Impaired Water Homeostasis and Developmental Defects in <i>Bombyx mori</i>.

Sun Lili L Zhang Zhongjie Z Zhang Ru R Yu Ye Y Yang Fangying F Tan Anjiang A

Frontiers in physiology 20200520

Insect ion transport peptides (ITPs) are important regulators of many physiological processes and they exert their functions by interacting with their receptors (ITPRs). In the current study, we comprehensively investigated the physiological functions of ITPR in the lepidopteran model insect, the silkworm (<i>Bombyx mori</i>), using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 nuclease (Cas9) genome editing technique. Mutations in silkworm IT ...[more]

PMID: 32508668

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Project description:BACKGROUND:Wing discs of B. mori are transformed to pupal wings during the larva-to-pupa metamorphosis with dramatic morphological and structural changes. To understand these changes at a transcriptional level, RNA-seq of the wing discs from 6-day-old fifth instar larvae (L5D6), prepupae (PP) and pupae (P0) was performed. RESULTS:In total, 12,254 transcripts were obtained from the wing disc, out of which 5,287 were identified to be differentially expressed from L5D6 to PP and from PP to P0. The results of comprehensive analysis of RNA-seq data showed that during larvae-to-pupae metamorphosis, many genes of 20E signaling pathway were up-regulated and those of JH signaling pathway were down-regulated. Seventeen transcription factors were significantly up-regulated. Cuticle protein genes (especially wing cuticle protein genes), were most abundant and significantly up-regulated at P0 stage. Genes responsible for the degradation and de novo synthesis of chitin were significantly up-regulated. There were A and B two types of chitin synthases in B. mori, whereas only chitin synthase A was up-regulated. Both trehalose and D-fructose, which are precursors of chitin synthesis, were detected in the hemolymph of L5D6, PP and P0, suggesting de novo synthesis of chitin. However, most of the genes that are related to early wing disc differentiation were down-regulated. CONCLUSIONS:Extensive transcriptome and DGE profiling data of wing disc during metamorphosis of silkworm have been generated, which provided comprehensive gene expression information at the transcriptional level. These results implied that during the larva-to-pupa metamorphosis, pupal wing development and transition might be mainly controlled by 20E signaling in B. mori. The 17 up-regulated transcription factors might be involved in wing development. Chitin required for pupal wing development might be generated from both degradation of componential chitin and de novo synthesis. Chitin synthase A might be responsible for the chitin synthesis in the pupal wing, while both trehalose and D-fructose might contribute to the de novo synthesis of chitin during the formation of pupal wing.

Project description:BackgroundMicroRNAs (miRNAs) play crucial roles in various physiological processes through post-transcriptional regulation of gene expressions and are involved in development, metabolism, and many other important molecular mechanisms and cellular processes. The Bombyx mori genome sequence provides opportunities for a thorough survey for miRNAs as well as comparative analyses with other sequenced insect species.Methodology/principal findingsWe identified 114 non-redundant conserved miRNAs and 148 novel putative miRNAs from the B. mori genome with an elaborate computational protocol. We also sequenced 6,720 clones from 14 developmental stage-specific small RNA libraries in which we identified 35 unique miRNAs containing 21 conserved miRNAs (including 17 predicted miRNAs) and 14 novel miRNAs (including 11 predicted novel miRNAs). Among the 114 conserved miRNAs, we found six pairs of clusters evolutionarily conserved cross insect lineages. Our observations on length heterogeneity at 5' and/or 3' ends of nine miRNAs between cloned and predicted sequences, and three mature forms deriving from the same arm of putative pre-miRNAs suggest a mechanism by which miRNAs gain new functions. Analyzing development-related miRNAs expression at 14 developmental stages based on clone-sampling and stem-loop RT PCR, we discovered an unusual abundance of 33 sequences representing 12 different miRNAs and sharply fluctuated expression of miRNAs at larva-molting stage. The potential functions of several stage-biased miRNAs were also analyzed in combination with predicted target genes and silkworm's phenotypic traits; our results indicated that miRNAs may play key regulatory roles in specific developmental stages in the silkworm, such as ecdysis.Conclusions/significanceTaking a combined approach, we identified 118 conserved miRNAs and 151 novel miRNA candidates from the B. mori genome sequence. Our expression analyses by sampling miRNAs and real-time PCR over multiple developmental stages allowed us to pinpoint molting stages as hotspots of miRNA expression both in sorts and quantities. Based on the analysis of target genes, we hypothesized that miRNAs regulate development through a particular emphasis on complex stages rather than general regulatory mechanisms.

Dataset Information

Molecular Disruption of Ion Transport Peptide Receptor Results in Impaired Water Homeostasis and Developmental Defects in Bombyx mori.

Publications

Molecular Disruption of Ion Transport Peptide Receptor Results in Impaired Water Homeostasis and Developmental Defects in <i>Bombyx mori</i>.

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